Reference to any prior art in this specification is not, and should not be taken as, an acknowledgment or any form of suggestion that this prior art forms part of the common general knowledge in Australia or any other country.
The increasing sophistication of recombinant DNA technology is greatly facilitating research and development in a range of technological fields such as the medical, horticultural and agricultural industries. Of particular importance, is the exploitation of naturally occurring genetic mechanisms in, for example, pathogens, to induce useful phenotypic changes in cells, such as plant cells. This is particularly evident in the horticultural field in relation to mechanisms to induce resistance in plants to a range of plant pathogens.
There has been considerable progress in the development of virus resistance in crops, for example, through transformation with transgenes derived from the target viruses. The most successful use of transgenes has been with RNA plant viruses. However, despite a number attempts to control single stranded DNA (ssDNA) plant viruses through transgenic resistance, strategies which have been successful for RNA plant viruses have not been as effective for ssDNA plant viruses. DNA plant viruses and particularly the single stranded DNA (ssDNA) plant viruses are responsible for significant commercial losses in a wide range of fruit, vegetable, grain and fibre crops in the tropics and sub-tropics.
There have been two groups of ssDNA viruses which infect plants. These are the Geminiviridae and the recently described nanovirus group. Members of the Geminiviridae have geminate virions and either a monopartite or bipartite circular ssDNA genome. Each molecule is about 2.7 kb in length. Of the Geminiviridae genera, the begomoviruses and the mastreviruses are the most important. The begomoviruses are whitefly-transmitted and have either monopartite or bipartite genomes. Members of this genus include some of the most economically devastating viruses of modem agriculture such as tomato (yellow) leaf curl (consisting of a range of different viruses spread through most tropical and sub-tropical regions), African cassaya mosaic (Africa), bean golden mosaic (South and Central America), mungbean yellow mosaic (India) and cotton leaf curl (South and South-East Asia) viruses. The impact of many of the begomoviruses has increased dramatically over recent years as a result of the widespread introduction of the aggressive “B biotype” of the whitefly vector, Bemesia tabaci. The mastreviruses have had a lesser impact on agriculture but are responsible for significant losses in some crops. These viruses are transmitted by the leafhoppers and have monopartite genomes. Members of this genus include maize streak (Africa), wheat dwarf (Europe) and tobacco yellow dwarf (Australia) viruses.
The nanoviruses have isometric virions and circular ssDNA genomes but these genomes are multi-component with at least six different integral genomic components each of which is approximately 1 kb. These viruses are transmitted by aphids except for one tentative nanovirus, coconut foliar decay virus, which is transmitted by a treehopper and has only been reported from Vanuatu. The economically most important nanovirus is banana bunchy top virus (BBTV) which nearly destroyed the Australian banana industry in the 1920s and causes major losses in the South Pacific, Asia and Africa. Subterranean clover stunt (Australia), faba bean necrotic yellows (Mediterranean) and coconut foliar decay (Vanuatu) viruses all cause significant yield loss.
The genome organization of the begomoviruses, the mastreviruses and the nanoviruses have significant differences including the number and size of genomic components and number and size of genes, the processing of transcripts, the orientation of genes and the like. There are, however, remarkable similarities which suggest that these viruses have very similar replication and infection strategies. All the gemini- and nanoviruses encode (i) a Rep protein which has nicking and joining activity and directs rolling circle replication of the viral genome; (ii) a virion coat protein; (iii) a protein that is involved in binding host cell retinoblastoma-like proteins resulting in the cell moving to S phase; (iv) a cell-to-cell movement protein; and (v) a nuclear shuttle protein. Further, the viruses have functionally similar intergenic regions (IR). For instance, the IR of begomovimses, the LIR of mastreviruses and the CR-SL of banana bunchy top nanovirus all contain (1) a stem/loop structure, the nonanucleotide loop sequence of which is highly conserved between all gemini- and nanoviruses and is the site of nicking and ligation by the Rep protein; and (ii) a domain within this region which recognizes the Rep protein. The SIR of the mastreviruses and the CR-M of banana bunchy top nanovirus are the site of binding of an endogenous primer responsible for priming the conversion of virion ssDNA into transcriptionally active dsDNA.
The success in developing transgenic resistance to RNA viruses in crops and the increasing demand for such resistance to ssDNA viruses has resulted in investigation of a wide range of strategies for ssDNA viruses targeting various viral genes including the coat protein gene, movement protein gene and the Rep protein gene. In addition, strategies using defective interfering DNAs and a suicide gene have been investigated. Most work in this area has involved begomoviruses rather than mastre- or nanoviruses.
In work leading up to the present invention, the inventors have exploited the replication mechanisms of ssDNA viruses in order to induce genetic resistance in plants. However, the present invention has wide ranging applications in modulating genetic activities such as expression of polynucleotide sequences to effect a particular phenotype in response to a stimulus.